[Molecular identification of heparin from pigs by allele-specific PCR (AS-PCR) and amplification refractory mutation system (ARMS)].

The aim of this study is to develop a convenient and effective method for the identification of heparin from pigs (include Sus scrofa domestica Brisson and Sus scrofa riukiuanus). Based on sequences of D-loop region of pigs and the other animals, two pairs of highly specific primers were designed for distinguishing heparin of pigs from other animals. The primers were employed to amplify D-loop region of DNA templates extracted from pig and seven other animal species that amounted to 49 samples. AS-PCR (allele-specific PCR) and ARMS (amplification refractory mutation system) were all suitable for fast identification of heparin from pig with anneal temperature at 54-56 degrees C in AS-PCR and with wider anneal temperature in ARMS,at 52-58 degrees C. An about 170 bp DNA fragments were amplified from separately pigs and whereas no DNA fragment was amplified from other samples under the same reaction condition.

[SNP marker and allele-specific diagnostic PCR for authenticating herbs of Perilla].

AIM: To authenticate all the varieties of Perilla (single-species genus), to analyze sequences of rDNA ITS regions and single nucleotide polymorphism (SNP) within them and based on these, to design allele-specific diagnostic PCR primers. METHODS: The rDNA ITS regions of the perilla varieties were sequenced and analyzed by Clustal X 1.8, MEGA 3.0. Allele-specific diagnostic PCR primers that can authenticate all the perilla varieties were designed based on SNPs loci. RESULTS: The length of rDNA ITS sequences of perilla varieties ranged from 612 to 615 bp in size, including ITS1 (230 -232 bp), 5.8S (179 bp) and ITS2 (203 -204 bp). The GC content is about 61.5% - 61.9%. There is not only SNPs in non-coding region ITS1 and ITS2 (ncSNP), but also three coding SNPs (cSNP) loci in the conservative region of 5.8S. All the SNPs have only two allele loci polymorphism. The cSNP in 5.8S is related to the morphology variation among the varieties. Allele-specific diagnostic PCR primers have been designed according to SNPs loci to authenticate accurately all the seeds and leaves of Perilla varieties. CONCLUSION: SNPs in rDNA ITS region can be used as an effective molecular markers to authenticate all the varieties of Perilla.

[Genetic diversity and molecular authentication of wild populations of Dendrobium officinale by RAPD].

AIM: Genetic diversity, relationship and molecular authentication of total 8 wild populations of Dendrobium officinale were investigated using RAPD markers. METHODS: 10 random decamer primers were screened for Random Amplified Polymophic DNA (RAPD) fragments. A DNA molecular dendrogram was established based on cluster analysis by UPGMA (unweighted pair-group method with arithmetic average), and the relationship of the wild populations were analyzed, and all the wild populations were authenticated. RESULTS: A total of 439 loci with an average of 43.9 loci per primer and 54.9 loci per population were amplified from 8 wild populations by 10 effective primers. In the total 104 amplified bands, 95 were polymorphic, corresponding to 91.35% genetic polymorphism. The genetic distances were 0. 590 to 0. 727, with an average of 0. 686. CONCLUSION: Distinct genetic differences and extensive genetic diversity were presented among the wild populations. RAPD markers were an informative and useful tool for the genetic diversity, evaluation and authentication of wild populations of Dendrobium officinale. Primer S412 could be used to authenticate 8 wild populations completely.